Abstract

Elevated extracellular adenosine in the tumor microenvironment generates an immunosuppressive niche that promotes tumor growth and metastasis that is mediated via A2A receptor (A2AR) on immune cells. CPI-444 is a potent, oral, selective A2AR antagonist that has been well tolerated in Ph 1/1b studies in non-oncology indications. CPI-444 is being investigated in a Ph 1b study in solid tumors as a single agent and in combination with the anti-PD-L1 antibody atezolizumab. In preclinical studies, daily treatment of the syngeneic mouse model MC38 with CPI-444 (1, 10, 100 mg/kg) led to dose-dependent inhibition of tumor growth, leading to tumor elimination in ∼30% of treated mice. Combining CPI-444 (100 mg/kg, qd, 14 days) with anti-PD-L1 (200 μg, 3qw, 4 doses) treatment in MC38 models synergistically inhibited tumor growth and eliminated tumors in 90% of treated mice. When cured mice were later re-challenged with MC38 cells, tumor growth was rejected in 100% of challenged mice, indicating that CPI-444 induced systemic anti-tumor immune memory. CD8+ T cell depletion abrogated the efficacy of CPI-444 treatment as a single agent as well as in combination with anti-PD-L1, demonstrating a role for CD8+ T cells in mediating primary and secondary immune responses. Anti-tumor efficacy of CPI-444 ± anti-PD-L1 was associated with increased CD8+ cell infiltration and activation in MC38 tumor tissues, and a corresponding rise in PD-1 expression on CD8+ T cells in the spleen. Additionally, levels of immune checkpoints were modulated by treatment with CPI-444, including GITR, OX40, and LAG3 on tumor infiltrating lymphocytes and circulating T cells, suggesting a broad role for adenosine mediated immunosuppression. Based on these results and others, we have initiated a Phase 1b clinical trial to examine safety, tolerability, biomarkers, and preliminary efficacy of CPI-444 as a single agent and in combination with the anti-PD-L1 antibody, atezolizumab, in patients with non-small cell lung, melanoma, renal, triple negative breast, and other (head and neck, colorectal [MSI-H], bladder) tumors. Peripheral blood and tumor biopsies are collected pre- and post-treatment for biomarker analysis of adenosine pathway expression and activity, immune cell activation and tumor infiltration, and mutational burden. To demonstrate CPI-444 blockade of A2AR, a functional signaling assay was developed. Peripheral blood samples from treated patients were stimulated ex vivo with the adenosine analog NECA and CREB phosphorylation was quantified using flow cytometry in B and T cells. Following 14 days of CPI-444 treatment, NECA signaling was robustly inhibited at the doses selected for the Ph1b study in both the CPI-444 single agent and combination cohorts. In summary, this shows that CPI-444 induces anti-tumor immunity in mouse tumor models and inhibits adenosine signaling in lymphocytes of treated humans. This is the first demonstration of immune modulation in cancer patients receiving an adenosine antagonist. Evaluation of patient clinical responses and additional biomarkers are ongoing.